Model Systems for Flavoenzyme Activity. Regulation of Flavin Recognition via Modulation of Receptor Hydrogen-Bond Donor-Acceptor Properties

Article abstract of DOI:10.1021/jo961877c

We have synthesized a new family of receptors for flavins based on 6-aryl-2,4-(acyldiamino)-s-triazines. In these synthetic hosts, systematic variation of the spatially remote substituents on the 6-aryl ring alters the hydrogen-bond-donating abilities of the amide functionality and the hydrogen-bond-accepting properties of the triazine N(3). This variation results in a strong modulation of the efficiency of flavin binding, with association constants for the receptor flavin complexes ranging over an 8-fold range.

Full text of DOI:10.1021/jo961877c

836
J . Org. Chem. 1997, 62, 836-839
Mod el System s for F la voen zym e Activity. Regu la tion of F la vin
Recogn ition via Mod u la tion of Recep tor Hyd r ogen -Bon d
Don or -Accep tor P r op er ties
Robert Deans,† Graeme Cooke,‡ and Vincent M. Rotello*,†
Department of Chemistry, University of Massachusetts, Amherst, Massachusetts 01003, and
School of Chemical Sciences, University of East Anglia, Norwich NR4 7TJ , U.K.
Received October 4, 1996X
We have synthesized a new family of receptors for flavins based on 6-aryl-2,4-(acyldiamino)-s-
triazines. In these synthetic hosts, systematic variation of the spatially remote substituents on
the 6-aryl ring alters the hydrogen-bond-donating abilities of the amide functionality and the
hydrogen-bond-accepting properties of the triazine N(3). This variation results in a strong
modulation of the efficiency of flavin binding, with association constants for the receptor flavin
complexes ranging over an 8-fold range.
An important feature of biomolecular catalysts is their
ability to modulate the energetics of the binding event.1
Modeling of this ability provides us with insight into the
origin and nature of intermolecular forces. Additionally,
improving our control over the recognition event allows
us to more effectively model biological systems where
recognition and function are directly correlated.2,3 Fi-
nally, our ability to modulate the recognition process
allows us to control the mechanical properties of materi-
als and tune the reactivity of catalysts.
In previous research,2,4 we have established the rec-
ognition of flavins by diacyldiaminopyridines through
hydrogen bonding.5 While binding energies in these
systems could be regulated to a certain extent through
choice of acyl group,2 the individual steric and electronic
causes for this modulation could not be individually
determined.
F igu r e 1. Receptor 1-flavin 3 complex, as predicted by the
AMBER forcefield.8
tronic properties of the triazine nucleus. Since the sterics
of the binding surface of these receptors are identical,
the electronic effects of the substituents can be deter-
mined independently.
To provide enhanced versatility in the design of
recognition-based sensors and devices, and gain a better
understanding of the factors that govern hydrogen-
bonding processes, we have designed a system where the
effects of hydrogen-bond-donor and -acceptor capabilities
are decoupled from steric considerations. These receptors
are based on the diaminotriazine nucleus6 and utilize
three-point hydrogen bonding between receptors 1 and
flavin 3 (Figure 1).7 In these receptors, variation of the
spatially distant aryl substituent modulates the elec-
Resu lts
Receptors 1a -h were synthesized from the corre-
sponding nitriles via reaction with dicyandiamide and
potassium hydroxide9 to provide the sparingly soluble (in
CDCl3) diaminotriazines 2a -h (Scheme 1). Acylation
with isobutyryl chloride then provided receptors 1a -h ,
which were readily soluble in CDCl3.
Dim er iza tion of Recep tor s 1. The 1H chemical
shifts of the amide protons of receptors 1a -h in CDCl3
show a strong concentration dependence. 1H NMR
titrations of these receptors show a steady downfield shift
of the amide protons, indicative of formation of a hydrogen-
bound complex (Figure 2). The curves obtained from
these titrations were fitted to dimerization isotherms to
obtain dimerization constants (Kdim) (Table 1).10 The
energies of dimerization of these systems shows a general
dependence on the electronic properties of the substitu-
ents, with electron-withdrawing groups diminishing rec-
ognition and electron-releasing groups enhancing bind-
ing. Due to the complexity of the system, a direct linear
† University of Massachusetts.
‡ University of East Anglia.
X Abstract published in Advance ACS Abstracts, J anuary 15, 1997.
(1) Fersht, A. Enzyme Structure and Mechanism, 2nd ed.; Free-
man: New York, 1985.
(2) Breinlinger, E.; Niemz, A.; Rotello, V. M. J . Am. Chem. Soc. 1995,
117, 5379-5380.
(3) For
a recent monograph on hydrogen bonding in biological
systems see: J effrey, G.; Saenger, W. Hydrogen Bonding in Biological
Structures; Springer Verlag: Berlin, 1991. For recent model studies
of hydrogen bonding in biomimetic systems see: Gallo, E.; Gellman,
S. J . Am. Chem. Soc. 1993, 115, 9774-9788. Zimmerman, S.; Murray,
T. Tetrahedron Lett. 1994, 4077-4080. Nowick, J .; Chen, J .; Noronha,
G. J . Am. Chem. Soc. 1993, 115, 7636-7644.
(4) Niemz, A; Rotello, V. M. J . Mol. Rec. 1996, 9, 158-162.
(5) For binding of flavins with diaminopyridines see: Shinkai, S.;
He, G. X.; Matsuda, T.; Hamilton, A.; Rosenzweig, H. Tetrahedron Lett.
1989, 5895-5898. Tamura, N.; Mitsui, K.; Nabeshima, T.; Yano, Y. J .
Chem. Soc., Perkin Trans. 2 1994, 2229-2237. For binding of diami-
nopyridines to other imides see: Park, T.; Schroeder, J .; Rebek, J . J .
Am. Chem. Soc. 1991, 113, 5125-5127. Muehldorf, A. V.; Van Engen,
D.; Warner, J . C.; Hamilton, A. D. J . Am. Chem. Soc. 1988, 110, 6561-
6562.
(7) For other examples of receptors based on diaminopyridine-imide
interactions see: Hamilton, A. D.; Van Engen, D. J . Am. Chem. Soc.
1987, 109, 5035. Muehldorf, A. V.; Van Engen, D.; Warner, J . C.;
Hamilton, A. D. J . Am. Chem. Soc. 1988, 110, 6561.
(8) The MacroModel 4.0 implementation of the AMBER forcefield
(AMBER*)was used: Mohamadi, F.; Richards, N.; Guioda, W.; Lis-
kamp, R.; Caufield, C.; Chang, G.; Hendrickson, T.; Still, W. J . Comput.
Chem. 1990, 11, 440.
(6) For a melamine-based flavin receptor see: Tamura, N.; Kajiki,
T.; Nabeshima, T.; Yano, Y. J . Chem. Soc., Chem. Commun. 1994, 2583.
(9) Simons, J .; Saxton, M. Organic Syntheses; Wiley: New York,
1963; Collect. Vol. IV, p 78.
S0022-3263(96)01877-4 CCC: $14.00 © 1997 American Chemical Society

Model Systems for Flavoenzyme Activity
J . Org. Chem., Vol. 62, No. 4, 1997 837
Sch em e 1
F igu r e 3. Plot of dimerization energy versus Σσm,p.
.
F igu r e 4. Chemical shift changes of flavin 3 H(3) upon
addition of selected receptors 1.
F igu r e 2. Plot of the chemical shifts of the amide protons of
1a , 1d , 1g, and 1h as a function of concentration.
resulted in a smooth downfield shift in the resonance of
H(3) of flavin 3 (Figure 4). The resulting curve was fitted
to a 1:1 binding isotherm, with explicit compensation
made for the reduction in free receptor 1 due to dimer-
ization of 1 (Table 1).12 From this fit, both the association
constants and limiting shift values for the receptors 1a -
h -flavin 3 complexes were determined (Table 1). These
association constants ranged from 12 to 97 M-1, in
contrast to a recent study using alkyl(diacylamino)-
triazines that had association constants of approximately
Ta ble 1. Bin d in g Con sta n ts, En er getics, a n d Lim itin g
Sh ift Va lu es for Recep tor 1-F la vin 3 Com p lexes
a-c
c
a,c,d
c
a,d,e
Kdim
host (M-1
∆Gdim
(kcal/mol)
Ka
(M-1
∆Ga
δmax
)
)
(kcal/mol) (H(3))(ppm)
1a 95 ( 3 -2.68 ( 0.02 24 ( 4 -1.87 ( 0.09 16.4 ( 1.0
1b 75 ( 2 -2.54 ( 0.02 53 ( 10 -2.34 ( 0.03 12.2 ( 0.04
1c 41 ( 2 -2.18 ( 0.05 97 ( 4 -2.69 ( 0.02 13.3 ( 0.1
1d 63 ( 2 -2.44 ( 0.02 63 ( 5 -2.44 ( 0.04 12.9 ( 0.2
1e 50 ( 3 -2.27 ( 0.03 58 ( 3 -2.36 ( 0.03 11.9 ( 0.2
1f
88 ( 2 -2.63 ( 0.02 44 ( 3 -2.23 ( 0.04 13.1 ( 0.3
6 M-1 13
It was proposed that these low observed binding
.
1g 60 ( 2 -2.41 ( 0.02 20 ( 2 -1.76 ( 0.06 14.8 ( 0.6
1h 80 ( 3 -2.58 ( 0.02 12 ( 4 -1.46 ( 0.16 19.8 ( 2.0
constants were due to the acyl groups of the triazine
a
b
CDCl3, 23 °C. Amide peak followed. c Errors represent the
d
standard error of the data fit to the calculated isotherm. H(3)
(10) Dimerization constants (Kd) for receptors 1a -h were obtained
via NMR titration, with the shift of the amide proton followed as a
function of concentration. Conners, K. Binding Constants; Wiley and
Sons: New York, 1987. To provide Kd, the data were fitted to the
equation
peak followed. e ppm downfield from TMS.
relationship between free energy and electron donor/
acceptor strengths would not be expected. A plot of
dimerization energies (-∆Gdim) versus Σσm,p (Figure 3)11
shows a strong correlation between the free energy of
dimerization and substituent donor/acceptor values, with
receptors 1a and 1d as the only significant outlying
points.
2
1/2
δd - δm
1
1
4Kd
δobs ) δm
+
[H] +
-
[H] +
- [H]2
(
)((
4K ) ((
)
) )
[H]
d
where the experimentally determined parameters are as follows: [H],
the total concentration of analyte, and δobs, the observed shift.
Parameters obtained through fitting are δm, the shift of the monomer,
δd, the shift of the dimer, and Kd, the dimerization constant.
(11) Chapman, N.; Shorter, J . Correlation Analysis in Chemistry;
Plenum: New York, 1978.
Recogn ition of F la vin 3 by Recep tor s 1. Addition
of receptors 1a -h to solutions of flavin 3 in CDCl3

838 J . Org. Chem., Vol. 62, No. 4, 1997
Deans et al.
F igu r e 6. Plot of the binding energies vs Σσm,p for the
complexes of receptor 1-flavin 3.
F igu r e 5. Representation of the two predicted conformations
adopted by the acyl groups in the triazine 1-flavin 3 complex.
observed for 1f and 1b, the two receptors with electron-
releasing substituents para to the triazine ring.14
The observed relationship between binding efficiency
and electron-donor/-acceptor properties of the aromatic
substituents can be attributed to concomitant changes
in the electrostatic potential and polarizabilites of the
hydrogen-bonding surface.15 The hydrogen-bonding sur-
face of receptors 1 can be divided into two components:
the hydrogen-bond-donating amides and the hydrogen-
bond-accepting N(3) position of the triazine. Changing
the electron density of the triazine nucleus affects both
of these recognition elements. As the electron-withdraw-
ing abilities of the substituents on the aromatic ring
increase, electron density in the triazine ring of receptors
1 decreases, increasing the positive electrostatic potential
of the amide protons. This increases the hydrogen-bond-
donating ability of the host, enhancing recognition.16
Concurrently, decreased electron density enhances the
negative potential and, hence, basicity of the triazine ring
nitrogen. This is shown in the decrease in the maximal
shift values (δmax) for the flavin 3 H(3) observed with
receptors featuring electron-withdrawing substituents.
While this decrease in basicity should result in energeti-
cally less favorable hydrogen bonding,17 the diminished
strength of the single hydrogen bond acceptor is overcome
by the enhancement of the two other hydrogen-bond
donors, providing the overall trend observed.
adopting a cis configuration (ADADA), creating ad-
ditional repulsive interactions during the binding event.
The enhanced binding observed in our system is most
likely caused by unfavorable steric interactions between
the aryl group and the side chains of the acyl groups in
the cis configuration, resulting in the preferred DAD
configuration required for complexation (Figure 5).
As shown in Table 1, variation of the substituents on
the aromatic ring of receptors 1a -h markedly influences
the efficiency of flavin 3 recognition. Since receptors
1a -h present sterically identical hydrogen bonding
surfaces for the recognition of flavin 3, all changes in
binding energies in receptor-flavin complexes can be
directly attributed to changes in the electrostatic poten-
tials and polarizability of the receptor hydrogen bonding
surface. Consequently, the efficiencies of binding in the
receptor 1-flavin 3 complexes are determined directly
by the nature of the substituents on the phenyl ring:
electron-withdrawing substituents enhance binding and
electron-donating groups diminish binding efficiency. The
correlation between recognition and the electron-donat-
ing/-withdrawing properties of the substituents is not
direct: a plot of binding energy (-∆Ga) versus Σσm,p
(Figure 6) indicates a generally linear correlation for the
bulk of the systems studied, with strong deviations
In summary, we have developed a family of diamino-
triazine-based receptors for flavin derivatives. These
receptors modulate the recognition of flavin derivatives
through variation of the electronic characteristics of the
hydrogen bonding surface. These receptors show a direct
correlation between the electronic properties of the
receptor and the efficiency of recognition, allowing fine
(12) Explicit solution of the simultaneous equilibria for host-guest
and guest-guest binding leads to a cubic expression. To provide Ka
and δHG, the data were fitted to the equation
3
2
δobs - δH
δHG - δH
2Kd
δobs - δH
1
[Gt] ) [Ht]
+
- [Gt] - 2[Ht] -
+
2
(
)
(
K )(δ
)
- δH
HG
Ka
a
δobs - δH
1
2[Gt] + [Ht] +
(14) The large difference in binding constants between the p-
(
K )(δ
)
- δH
a
HG
methoxy receptor 1f and the m-methoxy receptor 1g indicate
a
where the experimentally determined parameters are as follows: [Gt]
and [Ht], the total guest and host concentrations, respectively, δobs the
observed shift, δH, the shift of the host in the absence of guest, and
Kd, the guest dimerization constant. Parameters determined through
fitting are Ka, the host-guest association constant, and δHG, the
chemical shift of the host-guest complex.
substantial resonance contribution to the modulation of flavin recogni-
tion in these receptors. Due to the complex additivity of the various
effects, further analysis was not performed.
(15) For a study on the effects of electron-donating and -accepting
groups in urea and thiourea binding of sulfonic acids see: Wilcox, C.;
Kim, E.; Romano, D.; Kuo, L.; Burt, A.; Curran, D. Tetrahedron 1995,
621-634.
(16) Tsai, R. S.; Fan, W.; Tayar, N.; Carrupt, P. A.; Testa, B.; Kier,
L. J . Am. Chem. Soc. 1993, 115, 9632-9639. Chen, C. T.; Siegel, J . J .
Am. Chem. Soc. 1994, 116, 5959-5960.
(17) Neder, K.; Whitlock, H. J . Am. Chem. Soc. 1990, 112, 9412-
9413.
(13) Beijer, F. H.; Sijbesma, R. P.; Vekemans, J . A. J . M.; Meijer, E.
W.; Kooijiman, H.; Spek, A. L. J . Org. Chem. 1996, 61, 6371-6380.
Also useful are: Park, T. K.; Feng, Q.; Rebek, J . J . Am. Chem. Soc.
1992, 114, 4529-4532. Willner, I; Rosengaus, J ; Biali, S. Tetrahedron
Lett. 1992, 3805-3808. Simanek, E. E.; Wazeer, M. I. M.; Mathias, J .
P.; Whitesides, G. M. J . Org. Chem. 1994, 59, 4904-4909.

Model Systems for Flavoenzyme Activity
J . Org. Chem., Vol. 62, No. 4, 1997 839
Hz), 8.57 (2H, d, J ) 7.9 Hz), 8.81 (2H, br s). Anal. Calcd for
C18H20N5O2F3: C, 54.66; H, 5.1; N, 17.72. Found: C, 54.77;
H, 5.14; N, 17.52. 1e (33%): mp 235.5-236 °C; FT-IR (KBr)
3256, 3185, 2969, 1690 cm-1; 1H NMR (CDCl3) δ 1.34 (12H, d,
J ) 6.9 Hz), 3.36 (2H, septet, J ) 6.9 Hz), 7.66 (1H, t, J ) 7.9
Hz), 7.85 (1H, d, J ) 7.2 Hz), 8.69 (1H, s), 8.71 (1H, d, J ) 6.5
Hz), 9.31 (2H, br s). Anal. Calcd for C18H20N5O2F3: C, 54.66;
H, 5.1; N, 17.72. Found: C, 54.73; H, 5.24; N, 17.92. 1f
(26%): mp 224.5-225 °C; FT-IR (KBr) 3256, 3174, 2969, 2836,
1679 cm-1; 1H NMR (CDCl3) δ 1.32 (12H, d, J ) 6.9 Hz), 3.41
(2H, septet, J ) 6.9 Hz), 3.90 (3H, s), 6.99 (2H, d, J ) 9.0 Hz),
8.42 (2H, d, J ) 9.0 Hz), 9.24 (2H, br s). Anal. Calcd for
C18H23N5O3: C, 60.47; H, 6.49; N, 19.6. Found: C, 60.25; H,
6.37; N, 19.60. 1g (21%): mp 209-210 °C; FT-IR (KBr) 3246,
tuning of the binding event. Applications of these
receptors for the modulation of flavin reactivity are
underway and will be reported in due course.
Exp er im en ta l Section
Gen er a l Meth od s. Chemicals were purchased from Acros
Organics, Aldrich, Eastman Organic Chemicals, and EM
Science and used as received. Thin layer chromatography
(TLC) and column chromatography were carried out on glass
precoated TLC plates with silica gel 60 and silica gel 60 (230-
400 mesh), respectively. All reactions were performed under
an argon atmosphere. Microanalyses were performed by the
University of Massachusetts (Amherst) Microanalysis Service.
Fourier transform infrared spectra (FTIR) were measured on
a Perkin-Elmer Model 1600 FT-IR spectrophotometer. 1H
NMR spectra were recorded on a Bruker/IBM AC200 (200MHz)
spectrometer. All spectra were recorded using either CDCl3
or DMSO-d6 as solvent.
1
3174, 2826, 1679 cm-1; H NMR (CDCl3) δ 1.33 (12H, d, J )
6.9 Hz), 3.47 (2H, septet, J ) 6.9 Hz), 3.91 (3H, s), 7.15 (1H,
ddd, J ) 7.9, 2.5, 1.1 Hz), 7.42 (1H, t, J ) 7.9 Hz), 7.98 (1H,
app dd, J ) 2.5, 1.1 Hz), 8.09 (1H, app dt, J ) 7.9, 1.1 Hz),
9.47 (2H, br s). Anal. Calcd for C18H23N5O3: C, 60.47; H, 6.49;
N, 19.6. Found: C, 60.18; H, 6.38; N, 19.79. 1h (82%): mp
Gen er a l P r oced u r e for th e P r ep a r a tion of th e Dia m i-
n otr ia zin es 2a -h . To a solution of potassium hydroxide (168
mg, 3 mmol) in 1-pentanol (10 mL) were added dicyandiamide
(1.51 g, 18 mmol) and the starting nitrile (15 mmol). The
reaction mixture was then stirred for 24 h at 140 °C. After
cooling, the resulting solid was suspended in boiling water,
filtered, and dried. The diaminotriazines 2a -h thus obtained
were found to be sparingly soluble in CDCl3 but showed
satisfactory NMR spectra in DMSO-d6 and were thus used
without further purification. The yields obtained for the
various diaminotriazines were as follows: 2a (78%); 2b (75%);
2c (95%); 2d (77%); 2e (53%); 2f (97%); 2g (99%); 2h (99%).
Gen er a l P r oced u r e for th e P r ep a r a tion of th e Acyl-
a ted Dia m in otr ia zin es 1a -h . To a suspension of the
triazine (5 mmol) in pyridine (10 mL) was added dropwise
isobutyryl chloride (2.62 mL, 25 mmol) at room temperature.
The reaction mixture was then stirred at 100 °C for 24 h, after
which time the pyridine was removed under a stream of air.
The resulting solid was dissolved in CH2Cl2 (25 mL), washed
with a saturated aqueous solution of NaHCO3 (25 mL) and
then H2O (25 mL), and dried (Na2SO4) and the CH2Cl2
evaporated under reduced pressure. The resulting solid was
purified by column chromatography on silica gel with 5:1
hexanes/ethyl acetate and recrystallized from MeOH. 1a
(83%): mp 211.5-212.5 °C; FT-IR (KBr) 3256, 3185, 2964,
1685 cm-1; 1H NMR (CDCl3) δ 1.33 (12H, d, J ) 6.9 Hz), 3.44
(2H, septet, J ) 6.9 Hz), 7.47-7.64 (3H, m), 8.47 (2H, app dt,
J ) 6.9, 1.5 Hz), 9.33 (2H, br s). Anal. Calcd for C17H21N5O2:
C, 62.35; H, 6.47; N, 21.4. Found: C, 62.31; H, 6.36; N, 21.09.
1b (53%): mp 213-213.5 °C; FT-IR (KBr) 3246, 3185, 2969,
1685 cm-1; 1H NMR (CDCl3) δ 1.32 (12H, d, J ) 6.9 Hz), 2.49
(3H, s), 3.44 (2H, septet, J ) 6.9 Hz), 7.31 (2H, d, J ) 7.9 Hz),
8.36 (2H, d, J ) 8.3 Hz), 9.47 (2H, br s). Anal. Calcd for
C18H23N5O2: C, 63.31; H, 6.79; N, 20.52. Found: C, 63.09; H,
6.72; N, 20.72. 1c (71%): mp 238-239 °C; FT-IR (KBr) 3246,
232-233 °C; FT-IR (KBr) 3226, 2969, 2826, 1685 cm-1 1H
;
NMR (CDCl3) δ 1.32 (12H, d, J ) 6.9 Hz), 3.41 (2H, septet, J
) 6.9 Hz), 3.98 (3H, s), 4.01 (3H, s), 6.97 (1H, d, J ) 8.7 Hz),
7.99 (1H, d, J ) 1.8 Hz), 8.14 (1H, dd, J ) 8.7, 1.8 Hz), 9.04
(2H, br s). Anal. Calcd for C19H25N5O4: C, 58.89; H, 6.51; N,
18.08. Found: C, 58.94; H, 6.45; N, 17.69.
1H NMR Titr a tion s. Dim er iza tion of Recep tor 1.
Complexation studies were performed in CDCl3, a non-
competitive solvent, to allow the observation of specific hy-
drogen bonds. The dimerization constants (Kd) were obtained
by means of NMR concentration studies, carried out on the
receptors 1a -h . With the exception of receptors 1b, 1e, and
1f, this involved addition of aliquots of 0.1 M receptor solution
to an NMR tube containing CDCl3. The receptor concentration
ranged from 0.001 96 M (initially) to 0.075 M (finally). For
receptors 1b, 1e, and 1f, due to solubility problems, aliquots
of 0.05 M receptor solution were added. The receptor concen-
tration ranged from 0.000 98 M (initially) to 0.0375 M (finally).
Recep tor 1-F la vin 3 Bin d in g. To a host solution of flavin
3 (5 × 10-3 M) were added aliquots of receptor guest solution
1a -h (5 × 10-2 M). This resulted in a smooth downfield shift
in the resonance of H(3) of flavin 3. Final concentrations:
[Guest]total ) 0.025 M; [Host]total ) 0.0025 M. Nonlinear least-
squares curve fitting was performed, with the resulting curve
providing a good fit to a 1:1 binding isotherm, when compensa-
tion had been made for the dimerization of the receptor 1a -
h . From this curve fit it was possible to determine both the
association constant (Ka) and the limiting shift value (δmaxH(3))
for the various receptor 1a -h -flavin 3 complexes.
Ack n ow led gm en t. This research was supported by
the National Science Foundation (CHE-9528099), the
Petroleum Research Fund, administered by the Ameri-
can Chemical Society (30199-G4), and the NATO Col-
laborative Research Program (SRG-940781). V.R. thanks
Research Corporation for a Cottrell Fellowship.
1
3185, 2969, 1685 cm-1; H NMR (CDCl3) δ 1.33 (12H, d, J )
6.9 Hz), 3.23 (2H, septet, J ) 6.9 Hz), 8.35 (2H, d, J ) 8.7
Hz), 8.65 (2H, d, J ) 8.7 Hz), 9.02 (2H, br s). Anal. Calcd for
C17H20N6O4: C, 54.82; H, 5.42; N, 22.58. Found: C, 54.74; H,
5.83; N, 18.50.18 1d (92%): mp 210.5-211.5 °C; FT-IR (KBr)
3256, 3185, 2969, 1690 cm-1; 1H NMR (CDCl3) δ 1.32 (12H, d,
J ) 6.9 Hz), 3.27 (2H, septet, J ) 6.9 Hz), 7.77 (2H, d, J ) 8.3
Su p p or tin g In for m a tion Ava ila ble: Titration graphs
(dimerization and association) for receptors 1a -h . NMR
spectra of diaminotriazines 2a -h and 1a -h and IR spectra
of receptors 1a -h (32 pages). This material is contained in
libraries on microfiche, immediately follows this article in the
microfilm version of the journal, and can be ordered from the
ACS; see any current masthead page for ordering information.
(18) The elemental analysis results for receptor 1c always showed
low nitrogen content. This was attributed to the fact that one of the
nitrogen atoms had directly attached oxygens.
J O961877C